Species identification in dairy products by three different DNA-based techniques

Abstract

Polymerase chain reaction (PCR) with primers encoding a partial sequence of the β-casein gene was performed to detect the corresponding DNA in milk and cheese after an adapted DNA extraction procedure. In the PCR product from ovine or caprine β-casein DNA was shown to contain a specific restriction enzyme site that is not present in bovine β-casein DNA. Accordingly, after selected restriction enzyme analysis and horizontal polyacrylamide gel electrophoresis (PAGE), the undigested bovine β-casein fragment can be detected as an additional band if cow’s milk is present. Appropriate experiments using unprocessed milk demonstrated that a semi-quantitative assay could be established. The detection limit was about 0.5% cow’s milk in ewe’s and goat’s milk cheese. By use of a DNA intercalating agent the β-casein PCR products from cow or buffalo could be distinguished from those of ewe or goat as a consequence of sequence-specific retardation during agarose gel electrophoresis. Furthermore, single-stranded conformation polymorphism (SSCP) analysis was applied to detect expected species-specific conformation of the selected β-casein DNA sequences from the milk of cows, ewes, goats and buffalos milk. These techniques are compared with respect to their special use and application.